1. Field of the Invention
The present invention relates to a method of diagnosing mammalian tumors and more particularly to a photoidentification method of diagnosing tumor tissue using a synthetic carotenoporphyrin which consists of a carotenoid polyene covalently linked to a porphyrin or related cyclic tetrapyrrole. The diagnostic agents preferentially localize in tumor tissue and fluoresce upon exposure to light. An improved method of synthesizing the diagnostic agents employed herein is also provided.
2. Prior Art
Carotenoid pigments, ubiquitous in photosynthetic membranes, are essential for the survival of green plants. Two facets of carotenoid function are recognized in photosynthetic membranes. First, carotenoids photoprotect by rapidly quenching chlorophyll triplet states which are formed in antenna systems or photosynthetic reaction centers. This triplet-triplet energy transfer prevents chlorophyll-photosensitized formation of highly destructive singlet oxygen which is injurious to the organism. In addition, carotenoids act as antennas by absorbing light in spectral regions where chlorophyll absorbs weakly and delivering the resulting exitation to chlorophyll via a singlet-singlet energy transfer process. Finally, nearby carotenoids quench chlorophyll first excited singlet states. This quenching has been ascribed to energy transfer or electron transfer or some other process leading to internal conversion and is believed to play a role in the regulation of photosynthesis.
A number of porphyrin materials have been found to localize in tumor tissue and to damage that tissue upon irradiation with light. Many of these are being investigated as therapeutic agents ("hematoporphyrin derivative" and related materials). All of these agents suffer from the problem that they are also absorbed by healthy tissue which is also harmed by the light.
Various synthetic carotenoids designed to mimic carotenoid photoprotection have been investigated by researchers at Arizona State University. Synthetic carotenoporphyrins consisting of a carotenoid part covalently linked to a synthetic meso-tetraarylporphyrin which successfully exhibited both the photophysical functions of cartenoids in photosynthesis were first reported by Gary Dirks, Ana L. Moore, Thomas A. Moore and Devens Gust in Photochemistry and Photobiology, Vol. 32, pp 277-280 (Permagon Press Ltd. Great Britain, 1980).
A carotenoporphyrin which demonstrated quenching of the porphyrin triplet state by the attached carotenoid via triplet-triplet energy transfer was reported by R. V. Bensasson, E. J. Land, A. L. Moore, R. L. Crouch, G. Dirks, T. A. Moore and D. Gust in Nature, Vol. 290, No. 5804, pp 329-332, Mar. 16, 1981.
Since that time, various compounds which exhibit the triplet-triplet energy transfer described in Nature, supra, have been reported by the Arizona State University group.
In 1984, carotenoporphyrins 1-5 were prepared by Dr. Paul Liddell at Arizona State University and reported in his doctoral thesis dated December, 1985. Carotenoporphyrins 1-3 were reported by Harry A. Frank, Barry W. Chadwick, Jung Jin Oh, Devens Gust et al., Biochemical et Biophysical Acta 892 (1987) 253-263.
It has now been discovered that certain synthetic carotenoporphyrins preferentially localize in mammalian tumor tissue where they absorb and emit light when irradiated with light so that the site of the tumor may be detected by the fluorescence of the localized carotenoporphyrin. Localization of the carotenoporphyrins employed in the practice of the present invention is better than that of porphyrins alone and most importantly, photodamage of tissue is precluded by the quenching of the porphyrin triplet state. Thus the present invention overcomes the problem inherent with the existing porphyrin photosensitizing compounds as diagnostic agents, that of collateral tissue damage. In fact, with the prior art porphyrins, the entire body of the mammalian host becomes photosensitive, and exposure of any body parts to light must be avoided for from several weeks to months after treatment.